Lecture 30

Question 1

How are heavy and light chain gene families encoded

Answer 1

On separate chromosomes

Question 2

What is somatic recombination

Answer 2

The process by which B cell segments are rearranged to create different possible antibodies
Tightly regulated
Many of the same proteins involved in DNA repair

Question 3

Genes in Light chain

Answer 3

V, J, C
Variable region, Joining region, and Constant region genes

Question 4

Genes in Heavy chain

Answer 4

V, D, J, C
Variable region, Diversity region, Joining region, Constant region gene
D–>only in Heavy chains

Question 5

Where are CDR 1 and 2 encoded

Answer 5

In V segments of light and heavy chains
Both V regions are in germline DNA

Question 6

Where is CDR3 encoded in heavy and light chains

Answer 6

Heavy chain–>V, D, J gene segment
Light chain–> V-J segment

Question 7

Types of light chains

Answer 7

Kappa and lambda chains
Only 1 chains will be expressed while silencing the other
2 Loci

Question 8

How many loci in Heavy chain

Answer 8

1
Different constant regions represent different isotypes

Question 9

Overall pathway for somatic recombination

Answer 9

Inherited stem cell (B cell precursor)–>somatic recombination–>recombinase proteins join separate gene segments together–>mature naive B cell
Irreversible

Question 10

How many V regions to chose from

Answer 10

46

Question 11

How many D regions to chose from

Answer 11

23

Question 12

How many J regions to chose from

Answer 12

6

Question 13

What is combinatorial diversity

Answer 13

Diversity that results in different combinations of V, D, and J regions

Question 14

What do Recombination signal sequences (RSSs) do

Answer 14

RSSs flank each antibody gene segment to then be recognized be Recombinase enzymes

Question 15

What does each Recombination signal sequence (RSS) contain

Answer 15

Nonamer (9bp) and heptamer (7bp) sequence with either a 12 OR 23 bp spacer

Question 16

What is the 12/23 Rule

Answer 16

That 12bp RSS MUST pair with a 23bp RSS for recombination to occur

Question 17

What pairing does RSS direct in light and heavy chains

Answer 17

Light chain–> V-J
Heavy chain–> D-J and V-DJ

Question 18

What is within the Loop of DNA formed after RSS regions are brought together

Answer 18

Segments that were not selected
In loop–>out of segment

Question 19

Function of RAG-1 and RAG-2

Answer 19

RAG= recombination activating gene
Recognizing and cutting DNA at the immunoglobulin-encoding region and the RSS
Necessary for recombination

Question 20

Structure of RAG

Answer 20

Covalently closed DNA hairpin ends

Question 21

What is the signal joint

Answer 21

Loop of DNA excised when RSS are brought together
No longer on chromosome–>deleted

Question 22

What is the coding joint

Answer 22

Coding region of selected V and J regions that remain (not in loop)

Question 23

What is junctional diversity

Answer 23

When nucleotides are added or removed at junctions between V-D and D-J (or V-J for the light chain)
Occurs during recombination

Question 24

What is Artemis

Answer 24

Endonuclease that opens the DNA hairpins during junctional diversity

Question 25

Main mechanism of Junctional Diversity

Answer 25

Hairpin cleavage

Question 26

Ways that a hairpin can be opened (cleaved)

Answer 26

1) Exonuclease activity to remove nucleotides on each side of coding gene
2) Addition of palindromic (P) nucleotides at overhangs
3) Terminal Deoxynucleotidyl Transferase (TdT), can add up to 20 N-nucleotides (non-template encoded) to cleaved stranded mainly in heavy chain

Question 27

Function of repair enzymes in Junctional diversity

Answer 27

Trim off non-matching nucleotides,
Fill in remaining single-stranded gaps
Ligate new DNA